The sense of taste is important for determining food choice, for regulating food intake, and for ensuring efficient use of ingested nutrients. Taste can act as a warning system for the presence of potentially harmful foods, by, for example, the aversive sensations of sourness or bitterness, and as an attractant to potentially nutrient-rich foods, by, for example, the appealing sensations of sweetness, saltiness, and umami.
Taste stimuli are received by taste receptor cells assembled into taste buds that are located in the epithelium of taste papillae of the tongue (Kitagawa et al., Bioch. Bioph. Res. Comm., 283:236-242 (2001)). The stimuli are believed to be transduced by taste receptors at the surface of the taste receptor cells (Id.). The taste receptors encoded by the genes of a given species are reflective of that species' food choices. For example, the “sweet receptors” of an herbivorous species are expected to be different from those of a carnivorous species, since the two consume completely different diets whose foods contain different primary stimuli. Since taste receptor specificity likely reflects food choice, it follows that receptor sequence homology among species may be as predictive or more predictive of food preferences of a given species as phylogenetic relatedness among species.
Evolution has provided that each species' genes code for taste receptors unique to that species' food choices. For example, the “sweet receptors” of an herbivore are expected to be different from those of a carnivore, since the two consume completely different diets whose foods contain different primary stimuli. Even within the Order Carnivora, Feliformia (cat branch) and Caniformia (dog and bear branch) have different diets and show different taste responses to various sweeteners. Since taste receptor specificity must reflect food choice, it may follow that receptor sequence homology among species might be dependent more upon the types of foods consumed by individual species rather than by the phylogenetic relatedness of species. The behavior of carnivores, such as the domestic cat, towards stimuli such as sweet carbohydrates, which it cannot taste (Beauchamp, et al., J. Comp. Physiol. Psychol., 91(5):1118-1127 (1977)), and towards L-amino acids, which it can taste, should be explainable based on the specificity of the taste receptors of carnivores in general. The behavior of the domestic cat (Felis catus), a carnivore, towards stimuli such as sweet carbohydrates, which it generally cannot taste, and towards L-amino acids, which it generally can taste, should be explicable by the specificity of taste receptors of other carnivores.
The domestic dog and the domestic cat are two readily accessible and popular members of the Order Carnivora. Neurophysiological studies with dog show that it responds to chemicals representative of each of the five basic taste modalities: sweet, sour, bitter, salty, and umami. However, the spectrum of compounds within each taste group to which the dog responses are different from those to which the human, rodent, and cat respond (Bradshaw, Proc. Nutrition Soc., 50:99-106 (1991)). For example, while the dog responds to a range of mono- and d-saccharides and to some high intensity sweeteners, the cat does not. Particularly active in the dog are D-fructose, β-D-fructose, and sucrose. (Beauchamp et al., J. Comp. Physiol. Psychol., 91(5):1118-1127 (1977); Boudreau et al., Chem. Senses, 10:89-127 (1985); Boudreau (ed.), Neurophysiology and stimulus chemistry of mammalian taste systems. IN FLAVOR CHEMISTRY TRENDS AND DEVELOPMENTS. Washington D.C.: American Chemical Society (1989); Bartoshuk et al., Science, 171:699-701 (1971)).
Early studies suggest that domestic dog shows a preference for sucrose and that this behavior is congenital. (Grace & Russek, Physiology and Behavior, 4:553-558 (1968)). Additionally, domestic dog is believed to taste saccharin as bitter. (Grace & Russek, Physiology and Behavior, 4:553-558 (1968)). Electrophysiological studies showed that dog taste nerve fibers that responded to sucrose exhibited no response to saccharin and that the fibers fired by saccharin respond to the bitter alkaloid, strychnine (Anderson et al., Acta physiol scan, 21:105-119 (1950)). Experiments using amiloride show that the umami component of the canine chorda tympani nerve response is independent of the sodium component. (Kurihara & Kashiwayanagi, Ann. N Y. Acad. Sci., 855:393-397 (1998)). Direct knowledge of taste receptor genes of the domestic dog will allow insight into an animal's sensory world and may be useful for identifying modulators of the taste receptors encoded thereby to influence an animal's taste preferences.
Molecular receptors for the taste element of sweetness have recently been identified from human, mouse, and rat. Thus far, there are three known members of the T1R taste receptor family: T1R1, T1R2, and T1R3 (Montmayeur & Matsunami, Curr. Opin. Neurobiol., 12(4):366-371 (2002)). The T1R3 receptor gene is located within the Sac locus, the primary genetic locus controlling preference for sweet-tasting stimuli in mice (Li et al., Mamm. Genome, 12(1):13-16 (2001); Li et al., Mamm. Genome, 13(1):5-19 (2002)). The human syntenic region for the mouse T1R3 gene is on 1p36.33 (1162-1186 kb). The gene for T1R1 is located on human 1p36.23 (6324-6349 kb), which is ˜5 Mb from T1R3, and that for T1R2 is located on human 1p36.13 (18483-18729 kb), which is ˜12 Mb from T1R1.
Most of the T1Rs are G-protein coupled receptors with long N-terminal extracellular domains believed to be involved in ligand binding (Montmayeur & Matsunami, Curr. Opin. Neurobiol., 12(4):366-371 (2002)). Within the cell, the taste receptors heterodimerize, with T1R3 coupling separately with T1R1 and T1R2. In mouse, the T1R1/T1R3 heterodimer functions as a receptor for selected amino acids. The T1R2/T1R3 heterodimer functions as a receptor for stimuli considered sweet by humans. Current data indicate that the T1R3 component of the T1R heterodimer couples the taste receptor to cellular signal transduction processes, thereby ensuring that the stimulus-binding event is transduced to a neural signal. Thus, knowledge of the T1R receptors will lead to better understanding of species-specific reactions to sapid stimuli.
Currently, mechanisms for identifying novel taste stimuli for the domestic dog are limited, for example, to exhaustive and difficult feeding studies in which a novel ingredient is paired with a control ingredient and intake of the two are compared. Considerable time, effort, and expense can be expended in the discovery of a single stimulus. Furthermore, canine illnesses often are exacerbated by the animal's refusal to eat. Additionally, the molecular features that define acceptable taste stimuli for domestic dog remain largely unknown, making rational computational design approaches for taste stimuli difficult. As a result, knowledge of the canine taste receptor and its ligands may lead to a better understanding of dog taste perception and modulation thereof.
The present invention provides novel canine taste receptors, T1R1, T1R2, and T1R3, (also interchangeably referred to herein as Tas1r1, Tas1r2, and Tas1r3, respectively) methods of use thereof to identify compounds that can stimulate, inhibit, or modify the ingestive responses or general behavior of a dog. The screening methods of the invention allow the rapid screening of binding partners, agonists, antagonists, and modulators of the T1R receptors of the domestic dog. The results of the canine T1R receptor studies reflect the unique taste profile of the domestic dog.